This proposal is a response to an RFA from the NIAMDD (Dr. Sarah Kalser, Program Director). A novel validated growth-reinitiation assay using primary adult rat liver cell cultures shows that nanogram levels of known hepatotrophic factors promote hepatocyte proliferation under chemically defined conditions. We will use this system to follow the purification and the isolation of protein-like hepatocyte mitogens in blood or tissue extracts. Standard protein chemistry techniques are planned, including high pressure liquid chromatography. Tests of identify of purified factor(s) to known mitogens, and of specificity of purified factor(s) for non-liver cells will be performed. The effects of purified factor(s) on cellular processes required for hepatocyte proliferation--Na ion influx in early G1 and cAMP formation in late G1--will be studied. This work will broaden knowledge of mechanisms regulating liver regeneration. Basic information may lead to more rational approaches to treatment of acute and chronic liver disease.